April Journal-2009.p65 - Association of Biotechnology and Pharmacy

Current Trends in Biotechnology and Pharmacy
Vol. 3 (2) 188-196, April 2009. ISSN 0973-8916
receptor compartment by dialysis membrane with
molecular weight cut off between 12000 to 14000
(Himedia, Mumbai, India). The dialysis membrane
was previously soaked for 24 hours in PBS pH
7.4. The donor and receptor compartment hold
together using clamp. The receptor compartment
with 15 ml of PBS pH 7.4 was maintained at 37
± 0.5 o C and stirred with magnetic capsule
operated by magnetic stirrer, to prevent the
formation of concentrated drug solution layer
below the dialysis membrane. Samples of 3 ml,
were collected at predetermined time intervals
and replaced with fresh buffer. The concentration
of drug was determined by UV.
spectrophotometrically at 269 nm. Cumulative
percentage drug released were calculated (Table
3) and plotted against time (Fig. 1 and 2). The
data was fitted to different kinetic models to explain
the release mechanism and pattern using the
following equations.
Zero order equation Q = Q o
= Kt
First order equation Q = Q o
= e -Kt
Higuchi equation Q = Kt 1/2
Where, Q is the cumulative amount of
drug released, Q 0
is the initial amount of drug, k
is release constant and t is time.
Fig. 1. Schematic diagram of Franz diffusion cell
191
Preparation of Skin
Prior approval by Institutional Animal
Ethics Committee was obtained for conduction
of experiment (Ref: IAEC/SUCP/03/2007). The
albino rats were obtained from Sainath Animal
Agency, Hyderabad, India. Albino rats weighing
170-190 gm were sacrificed using anesthetic
ether. The hair of test animals was carefully
removed with the help of depilatory and the full
thickness skin was removed from the abdominal
region. The epidermis was prepared surgically
by heat separation technique (22), which involved
soaking the entire abdominal skin in water at 60
°C for 45 sec, followed by careful removal of the
epidermis. The epidermis was washed with water
and used for ex vivo permeability studies.
Ex vivo Skin Permeation Studies
The ex vivo skin permeation studies were
carried out using Franz diffusion cell (Fig. 1) with
a diffusional area of 3.73 cm 2 . Rat abdominal skin
was mounted between the compartments of the
diffusion cell with stratum corneum facing the
donor compartment. The receiver phase is 15 ml
of PBS pH 7.4, stirred at 300 rpm on a magnetic
stirrer. The stratum corneum side of the skin was
kept in intimate contact with the film and over
that placed a backing membrane. The whole
assembly was kept in a water bath at 37 ± 0.5
o
C. Samples (3 ml) were collected at
predetermined time intervals and replaced with
fresh buffer. The concentration of drug was
determined by U.V. spectrophotometrically at 269
nm. Cumulative percentage drug permeated was
calculated and plotted against time (Fig. 3 and 4).
Flux was determined directly as the slope of the
curve between the steady state values of the
amount of drug permeated (mg cm -2 ) v/s time
(hours) (23) and permeability coefficients were
deduced by dividing the flux by the initial drug
load (mg cm -2 ) as shown in Table 3.
Mamatha et al